Treatment of brain damage

ABSTRACT

The present invention relates to the treatment of brain damage by cellular transplantation. According to one aspect of the invention, a method for treating a motor, sensory and/or cognitive deficit comprises administering a composition comprising pluripotent cells into the damaged brain in a region contra-lateral to that containing the site of damage. The cells are preferably conditionally immortal.

FIELD OF THE INVENTION

This invention relates to the treatment of disorders associated withdamage to the brain. In particular, this invention relates to treatmentof disorders by cellular transplantation into a damaged brain.

BACKGROUND OF THE INVENTION

Stroke is the largest cause of adult disability worldwide. The incidenceof stroke is about 1.3% of the US population, and 39.4% of victims showsignificant residual impairments, ranging from hemiplegia to restrictedlimb use and speech defects. Approximately 60% of strokes are caused byocclusion of the middle cerebral artery (MCAo), resulting in damage inthe striatum and cortex with consequent deficits to sensory and motorsystems. There is therefore a substantial clinical need for treatmentsthat reduce or alleviate the deficits.

Typical therapies for stroke are aimed at interrupting the cascade ofevents that lead to intraneuronal calcium accumulation and cell death,and to provide stimulation through rehabilitation, e.g. physiotherapy,to promote intracerebral reorganisation. However, pharmacologicaltreatments must be administered quickly to protect against cell deaththat typically occurs within three hours of occlusion. In addition, thetherapy based on rehabilitation appears to be limited to a period of 3-6months after stroke, after which residual disabilities do not undergoappreciable reduction.

There has been much interest recently in the possibility oftransplanting new cells into the damaged neuronal system to promoterepair and alleviate the disorders. One difficulty associated with celltransplantation is the need to provide clonal cell lines from differentregions of the brain. This has proved to be a major difficulty inpreparing cells for transplantation. WO-A-97/10329 describes the use ofconditionally immortalised pluripotent neuroepithelial cells in thetransplantation into the damaged brain. The neuroepithelial cellsexpress a temperature-sensitive oncogene so that they are capable ofunlimited expansion under permissive low temperatures in vitro, butcease dividing to develop into mature neural cells on implantation intothe higher temperature of the brain (38° C). A particular advantage ofthese cells has been shown to be their ability to develop intosite-appropriate neurons or glia, under the control of signals from thehost brain, so that problems associated with choosing the correct tissuefor transplantation is avoided. It has also been shown that the cellscan migrate to the site of damage when transplanted into a regionproximal to the damaged site. Therefore, the use of these cells offers aviable alternative to pharmacological treatments for repair of braindamage.

However, although the cells were shown to migrate to discrete areas ofdamage, focal ischaemia results in extensive damage and it is by nomeans certain that areas of infarction would provide a sufficiently wellvascularised matrix to support the survival of grafted cells.

There is therefore the need for improvements in transplantation in orderto provide cells that successfully graft into the adult damaged brainand compensate for the deficits.

SUMMARY OF THE INVENTION

It has now been realised that pluripotent cells can successfully repairdamage when administered into the side of the brain contra-lateral tothat containing the site of damage.

Therefore, according to one aspect of the invention there is a methodfor treating brain damage comprising administering a compositioncomprising pluripotent cells into the damaged brain, whereinadministration is into the brain hemisphere contra-lateral to thatcontaining the site of damage.

Preferably, the pluripotent cells are neuroepithelial stem cells, inparticular, those from the MHP36 clonal cell line, defined herein.

The cells are preferably conditionally immortal. Immortalisation may beachieved by the transduction of a temperature-sensitive oncogene intothe cells as disclosed in WO-A-97/10329.

The advantage of administering the cells contra-laterally is that theintact (contra-lateral) region may provide a more tolerant environmentfor cell grafts, avoiding the inflammatory response at the site ofdamage which might cause cell rejection.

DESCRIPTION OF THE INVENTION

The cells of the present invention are capable of correcting a sensory,motor and/or cognitive deficit when implanted into the brain hemispherecontra-lateral to that of the damaged part of the human brain. The term“damage” used herein includes reduction or loss of function caused bycell loss. Damage may be caused by any of a variety of means includingphysical trauma, hypoxia (lack of oxygen), chemical agents, for example,damage may be caused by drug abuse, and disease. The following diseasesand pathological conditions are examples of diseases or conditions whichresult in deficits which may be treated in accordance with the presentinvention: traumatic brain injury, stroke, perinatal ischaemia,including cerebral palsy, Alzheimer's, Pick's and related dementingneurodegenerative diseases, multiple sclerosis, multi-infarct dementia,Parkinson's and Parkinson's-type diseases, Huntington's disease,Korsakoff's disease and Creuzfeld-Jacob disease. Amnesia, particularlyfollowing transitory global ischaemia such as after cardiac arrest orcoronary bypass surgery, may also be treated in accordance with thepresent invention.

The present invention is particularly suited to the treatment of strokewhere damage occurs primarily in one brain hemisphere e.g. due to anocclusion in the middle cerebral artery.

By “contra-lateral” it is intended that this refers to the hemisphere ofthe brain that does not contain the site of damage. Therefore, if thereis an occlusion in the left hemisphere, then, obviously, thecontra-lateral region is the right, undamaged, hemisphere.

Of course, in some instances damage may occur in both hemispheres, andin these cases the contra-lateral region is understood to be thehemisphere which exhibits least damage.

The term “pluripotent” is used herein to denote an undifferentiated cellthat has the potential to differentiate into different types ordifferent phenotypes of cell, in particular into cells having theappropriate phenotype for the intended use. The cell type or phenotypeinto which such a pluripotent cell finally differentiates is at leastpartly dependent on the conditions in which the cell exists or findsitself.

For use in the present invention the cells should be capable ofdifferentiating into cells appropriate to repair or compensate fordamage or disease in the target area of the brain. It will beappreciated that cells for transplantation need not be capable ofdifferentiating into all types or phenotypes of neural cells. The cellsmay, for example, be bipotent. However, a high degree of potency isgenerally preferred as this gives greater flexibility and potential fortransplantation into different areas of the brain.

Suitable pluripotent cells include those known in the art as “stemcells” and those called or known as “precursor cells”. In particular,neuroepithelial stem cells are suitable for use in the presentinvention. However, other cells may also be used. Alternative cells maybe those defined as haematopoietic stem cells which may be capable ofdifferentiating into neural cells.

The pluripotent neuroepithelial cells are advantageously, and willgenerally be, conditionally immortal and may be prepared as disclosed inWO-A-97/10329.

The treatment may be carried out on any mammal but the present inventionis especially concerned with the treatment of humans, especiallytreatment with human cells, and with human cells and cell lines.

To treat a patient it is necessary to establish where damage hasoccurred in the brain. This may be carried out by any method known inthe art, e.g. magnetic resonance imaging (MRI). Once the existence ofdamage has been established, whether it be in one isolated area or inseveral areas, treatment by implantation of cells into thecontra-lateral region to that of the damaged area may be carried out,again by conventional means. The pluripotent cells may be transplantedat a single site, or preferably at multiple sites, and may be able tomigrate to the site(s) of damage and, once there, differentiate inresponse to the local microenvironment, into the necessary phenotype orphenotypes to improve or restore function.

In addition to administering the cells into the contra-lateral region,it may also be desirable to co-administer the cells into the damagedhemisphere (ipsi-lateral region). Treatment in this manner may promotethe improvement or restoration of brain function by differentmechanisms.

Without wishing to be bound by theory, it may be that repair followingtransplantation into the contra-lateral region results from migration ofthe pluripotent cells into the area of damage, with the reconstitutionof local circuits to restore or sustain function. It may also be thatthe transplanted cells augment spontaneous processes within the intact(contra-lateral) side which attempt to compensate for the damage. If thelatter is correct, then it may be unnecessary for the transplanted cellsto cross to the side of damage to exert an effect.

It may be possible to promote repair by encouraging the activity ofparticular regions of the brain. By using passive or active exercise ofcertain regions, it may be possible to augment the spontaneous processesoccurring after transplantation. For example, it should be possible tostimulate particular brain regions by requiring certain tasks to beperformed. In doing so, the brain region may generate biological signalsthat aid repair.

The stimulation of the brain may be visualised using detectiontechniques such as magnetic resonance imaging (MRI). These techniquescan be adapted to permit the patient to visualise the active brainregions, so that, through the process of biofeedback, the patient canstimulate particular regions that may encourage repair.

Preferably, treatment will substantially correct a motor, sensory and/orcognitive deficit. However, that may not always be possible. Treatmentaccording to the present invention and with the cells, medicaments andpharmaceutical preparations of the invention, may lead to improvement infunction without complete correction. Such improvement will neverthelessbe worthwhile and of value.

The number of cells to be used will vary depending on the nature andextent of the damaged tissue. Typically, the number of cells used intransplantation will be in the range of about one hundred thousand toseveral million. Treatment need not be restricted to a singletransplant. Additional transplants may be carried out to further improvefunction.

Methods for transplantation of cells into humans and animals are knownto those in the art and are described in the literature in the art. Theterm “transplantation” used herein includes the transplantation of cellswhich have been grown in vitro, and may have been genetically modified,as well as the transplantation of material extracted from anotherorganism. Cells may be transplanted by implantation by means ofmicrosyringe infusion of a known quantity of cells in the target areawhere they would normally disperse around the injection site. Suitableexcipients and diluents will be apparent to the skilled person, based onformulations used in conventional cell transplantation.

The following non-limiting example illustrates the invention.

EXAMPLE

Conditionally immortal pluripotent neuroepithelial cells from the MHP 36clonal cell line were prepared as disclosed in WO-A-97/10329.

In the experiments described below, conditionally immortal cells usedfor transplantation are derived from the H-2K^(b)-tsA58 transgenic mousedeveloped by M. Noble and his associates at the Ludwig Institute forCancer Research (Jat et al., P.N.A.S. USA, 1991, 88:5096). All cellsfrom this mouse possess a temperature-sensitive oncogene (tsA58, thetemperature sensitive mutant of the SV40 large T antigen under thecontrol of the interferon-inducible H-2K^(b) promotor) such that thecells divide at the permissive temperature (lower than body temperature,33° C.) but differentiate only when restored to mouse body temperature(38° C.-39° C.). It is this feature that provides them with conditionalimmortality. This allowed us to clone and expand cell lines in vitrowhich then differentiated upon transplantation into a host brain. Anumber of cell lines were cloned from a population of cells takenoriginally from the transgenic mouse, specifically, from embryonic day14 (E 14) hippocampus. We studied the rats, which received transplantsof those cells, for at least 8 months and in no case did the cells,after transplantation, form tumors. Furthermore, in post-mortemhistological preparations, the transplanted cells (marked by priortransfection with a lac-z reporter gene) have the appearance ofdifferentiated cells appropriate to the rodent nervous system.

21 Wistar rats were subjected to left intraluminal occlusion of themiddle cerebral artery (MCAo-IL) under halothane anaesthesia asdisclosed in Ginsberg et al, Cerebrovascular disease, 1998; Volume1:14-35.

Following exposure of the left internal carotid artery, a 3.0 mm prolinefilament coated at the tip with silicon was inserted 18-20 mm up to thejunction of the circle of Willis and tied in place for 60 min.Anaesthesia was discontinued after insertion of the filament, and therat tested for neurologic deficit (contra-lateral paw flexion andcircling) to establish the presence of ischaemia. Anaesthesia wasresumed after 60 min for retraction of the filament to the externalcarotid stump, where it was left in place, the excess filament trimmedoff, and the wound sutured. Neurologic and health status were monitoredfor a week, until normal feeding was seen and post-operative weightregained. Control rats (n=11)were sham operated by exposure of the leftinternal carotid artery only.

Transplant and sham graft surgery was undertaken 2-3 weeks afterocclusion or sham surgery. Rats were anaesthetised with Immobilon (0.01ml/100 g, im) after pre-treatment with Hypnovel (midazolam: 0.03 ml/100,im), and placed in a stereotaxic frame. Holes were drilled in the rightside of the skull to allow the penetration of a 10 μl Hamilton syringeat the following coordinates (mm) derived from Bregma, with the skull inthe flat position (−3.2 mm). AP represents “anterior-posterior”, Lrepresents “lateral” and V represents “ventral”.

AP: −0.3 L: −3.5 V: −4.5, −6.0 L: −5.5 V: −4.0, −5.5 AP: −1.3 L: −3.0 V:−5.0, −6.5 L: −5.5 V: −5.0, −6.5

3.0 μl of suspension (25,000 cells/μl) were infused over 2 min at eachof the 8 sites, and the cannula was left in place for a further 2 min toallow diffusion from the tip. Medial descents were aimed for striatumand lateral descents were aimed for cortex at the estimated anterior andposterior extent of the area of infarct on the opposite side, to targetpotential regions of reorganisation. Controls received infusions ofvehicle.

Rats were tested from 4-6 weeks after transplantation over a period of10 months. During this time the performance on the repeated measure test(Bilateral Asymmetry Test) remained stable.

Bilateral Asymmetry Test

The bilateral asymmetry test (BAT) has been used to access a variety oflesions (Schallert et al, Pharmacol. Biochem. Behaviour, 1982;16:455-462). Strips of tape 1 cm wide and 5 cm long were wound roundeach of the two forepaws, in random order. Animals were placed in anobservation cage and timed for latency to contact and to remove eachtape. The side first contacted was also noted by the Experimenter blindas to the experimental groups. Random checks on reliability wereincluded by comparing the Experimenter's scores with those of a secondobserver; typically inter-rater reliability was above 90%. Rats weretested before surgery, and during the week before grafting, to establishpre- and post-operative baselines. One session of 4 tests of 3 min wascarried out each week, for 18 weeks, commencing 4-6 weeks aftertransplantation to assess long term recovery.

Rotation

This is a test for measuring rotational bias. When challenged withamphetamine, MCAO animals show a clear bias to the direction of thelesion.

Spontaneous and drug-induced rotation was measured approximately 38weeks after transplantation in an 8-bowl rotameter (TSE GmbH, BadHamburg) in which rats were harnessed for 30 min and swivels recorded ineither left or right direction. Rats were tested for response to saline(baseline). They were then tested once a week, on alternate weeks, witheither amphetamine (Sigma: 2.5 mg/kg) or apomorpine (Sigma: 0.5 mg/kg)on three occasions over a testing time of 6 weeks. All injections weregiven in a volume of 1.0 ml/kg, ip.

Before occlusion, rats did not show a mean difference in paw use asjudged by the latency to contact and removal of tapes from the left andright paw. After sham surgery the controls continued to show nopreference. However, rats subjected to MCAO showed a marked disparitybetween the two paws, with the right paw contacted and the tape removedsignificantly more slowly than the left, indicative of contra-lateralsensorimotor impairment. This robust and stable deficit persistedthroughout the 18 weeks of testing. In rats that received MHP36 grafts,the stroke-induced forepaw disparity was non-significant by 8 weeksafter transplantation, and this improvement persisted through the 18weeks of testing, so that there was no difference between paws. Hencegrafted rats did not differ from controls as both groups showed that thetwo paws were equivalent in latency to contact and remove the tape, andthat the right paw was contacted first as often as the left. In a groupof rats that were subjected to 60 min MCAO and which did not receivesham transplants, the paw disparity was comparable to that in the groupinjected on the intact side with a large volume of vehicle. This resultsuggests that injection damage on the intact side neither exacerbatednor reduced the extent of sensorimotor deficit induced by MCAO.

Baseline (spontaneous) rotation was mildly asymmetrical in stroke ratswithout grafts, in that there was more turning to the right than theleft, whereas control and grafted rats showed comparable turning in bothdirections. However, in response to amphetamine on weeks 2, 4, and 6 oftesting, stroke rats without grafts showed marked turning to the left,towards the lesioned side, indicative of asymmetric dopamine release onthe intact side. Group differences were very substantial and the nongrafted group differed significantly from the grafted and controlgroups, which did not differ in response to amphetamine. A similar, butless marked effect, was seen with the postsynaptic dopamine agonistapomorphine. Groups differed on weeks 3, 5 and 7 with the non-graftedstroke group showing more marked leftwards rotation than the grafted andcontrol groups which did not show a turning bias. In all groups thenumber of turns was lower in response to saline than to the dopamineagonists. However all groups showed a similar activation in response todrug, so that drug induced changes in bias in the non-grafted group werenot associated with differences in activity.

At the end of behavioural testing (approximately 11 monthspost-transplantation) histology studies were undertaken. Rats wereperfused with 4% paraformaldehyde, flushed through the upper bodyvasculature via a cannula inserted through the heart and into the aorta,which was attached to a pump. 50 μm coronal sections were cut throughthe brain, placed on gelatine-coated slides and frozen to preventdehydration. Serial sections were stained for the presence of β-Gallabelled cells, and cells reactive to glial fibrillary acidic protein(GFAP) and tyrosine hydroxylase (TH) to identify glial and neuronal celltypes within the graft.

In a first study, only one brain from the grafted and non-grafted groupssubjected to MCAO was examined. In both animals subjected to MCAO,cavitation was severe, amounting to approximately 75% of the hemispherevolume. Ventricles on the infarct side were enlarged, so that only athin strip of striatal tissue separated the lateral from the infarct.Ventricles were also enlarged to a lesser extent on the intact side, anddistortion, possible via oedema soon after occlusion, had pushed themidline towards the intact side. In the grafted animal β-Gal positivecells were seen at the injection site in the middle of the intactstriatum. Cells were also seen in a diagonal band stretching caudallyand laterally through the striatum towards the parietal cortex. β-Galpositive cells were seen approaching and within the side opposite toimplantation. They formed in a dense band along the lower ventricularmargin of the corpus callosum, and encircled the area of the infarct.They were particularly prominent in the residual strip of striatum, andsome had left the corpus callosum caudally to enter the cortex. Thesegrafted cells showed morphologies of several types, including bipolarcells, glial cells and neuronal cells of both pyramidal and medium spinyneuron appearance, suggesting a diverse pattern of differentiation.

For estimating lesion volume 50 μm sections were cut from 3.7 to 6.3 mmbefore bregma in all rats subjected to MCAO, with and without grafts,and intact controls. Every tenth section was collected giving aninter-section distance of 500 μm and a total of 20 sections per brain.Sections were strained with Cresyl Fast Violet. Images of each sectionwere taken using a stereo microscope and estimations of lesion volumewere carried out. For control rats there was no difference between thehemispheres. In rats subjected to 60 min of MCAO there was a substantialinfarct representing approximately 26% of the total brain volume. Instroke rats with MHP36 grafts, lesion volume was decreased toapproximately 16% of the total brain volume, and was significantlysmaller (p<0.05) than in rats with MCAO and sham grafts.

A later study compared the effect of grafts into either the ipsi-lateralor contra-lateral sides, or into the ventricles. Grafted cells werelabelled with the. fluorescent marker PKH26 to assist theiridentification. Behaviour was measured for 12 weeks aftertransplantation. Over this period there was a significant improvement inthe bilateral asymmetry test in rats with grafts in both theipsi-lateral and contra-lateral side, but not in those withintraventricular grafts. However, rats with intraventricular grafts,unlike those with intraparenchymal grafts, showed improved spatiallearning and memory in the water maze. These results indicate that thesite of grafting affects behavioural recovery and support the claim thatuse of multiple sites may be advantageous. In contrast to the finding inthe earlier study, where contra-lateral grafts restored spontaneous andamphetamine-induced rotation in animals tested 10 months aftertransplantation, there was no improvement in rotation bias in any of thegrafted groups tested 10-12 weeks after grafting. These findingssuggested that the time course of recovery may differ for differenttasks.

The brains of all the rats were processed for histology as describedabove. Grafted cells were visualised by PKH26 fluorescence, and doublelabelled with antibodies to neuronal and glial markers to identify cellsthat differentiated into these phenotypes. Site of implantationinfluenced cell survival and the pattern of migration. In general, morecells survived with grafts implanted contra-lateral to the lesion thaninto the ventricles, with ipsi-lateral grafts being intermediate.However, there was a similar proportion of neuronal cells, seenprimarily in the midline regions, in all granted groups. Importantly,grafts from all three sites migrated across the midline to the oppositeside of the brain. Thus about a third of grafted cells placed in thelesioned side were found in the intact side of the brain, whilst asimilar proportion of grafted cells also migrated from the intact to thelesioned side, as in the earlier study. These surprising findingsindicated that grafted cells not only responded to signals arising frominjury, but were also attracted to the intact side, possibly by signalsarising from processes of reorganisation.

Lesion volume, measured as described above, showed that the area ofdamage comprised approximately 18% of the total brain volume. Howeverthere was no difference between sham grafted animals with MCAO, andthose with grafts. Thus grafts did not significantly reduce lesionvolume, measured 14 weeks after transplantation, in contrast to thereduction seen at 11 months after transplantation in the earlier study.This may indicate that grafts give some protection against secondarydegeneration, and the effect is only clearly evident at a late timepoint.

In a later study, the remaining brains were sectioned and the lesionvolumes determined for each group by measuring the volumes of theipsi-lateral and contra-lateral hemisphere. Volume measurements of thelesion size revealed 18% loss of total brain volume in animals with 60minutes MCAO. No difference in lesion size was found between the MCAogroups regardless of transplantation site. When the total number oftransplanted cells was analysed according to implantation site, itemerged that grafted cells implanted contra-laterally were significantlygreater than those grafted in the ventricles, although there was nostatistical difference when compared to cells grafted ipsi-laterally. Itwas also apparent that there had been extensive migration away from theimplantation site into the opposite hemisphere.

The above experiments aimed to see whether grafts of MHP36 cells, from aconditionally immortalised clonal line, would promote functionalrecovery from stroke damage when placed in the intact hemispherecontra-lateral to the infarct cavity. The findings indicated that bothsensorimotor and motor asymmetries were normalised in rats with graftsinitially sited in the intact hemisphere.

The evidence for recovery of sensorimotor and motor functions is robust,because improvements were seen over an extended time period. Forexample, in MCA-occluded rats without grafts, tape-removal deficits wereseen consistently over an 18 week period of testing, with no hint ofspontaneous recovery. Improvement in grafted rats was also consistentover this period. The late rotational data is interesting in thatspontaneous deficits were manifest in mild rotation to the right,possibly reflecting a stronger push by the unaffected left paw, relativeto the right. However, dopamine agonist drugs induced marked rotation tothe left, consistent with activation of dopamine receptors on the right(intact) side of the brain. This asymmetry was not evident in rats withgrafts sited in the intact side, even though one might expect theasymmetry to be amplified, if grafted cells on the intact sidedifferentiated into tyrosine hydroxylase-positive (TH) neurons.

The control, grafted and non-grafted stroke groups showed a similardegree of locomotor stimulation in response to amphetamine (i.e. thenumber of rotations increased substantially above baseline) so that thedrug affected activity in all groups. If the normalisation of rotationbias in the grafted group reflected lesion-induced insensitivity todopamine stimulation on both sides of the brain, one would not expect tosee such a marked increase in rotation in response to amphetamine.Therefore it is reasonable to suppose that MHP36 grafts normalisedrotational behaviour by providing dopaminergic inervation on the side ofthe infarct. However, since this is a late effect, some othercompensatory mechanism, possibly involving reduced retrogradedegeneration, cannot be excluded.

There was some evidence of grafted cells on the side of implantation,not only around the sites of injection, but also forming a ventralstream of migration through the striatum. Thus it may be premature toconclude that grafted cells exert functional effects only if they crossto the side of damage. They may also be involved in reorganisation ofthe intact hemisphere. This conclusion is supported by finding thatcells implanted on the lesion side migrated to the intact contra-lateralside.

What is claimed is:
 1. A method for treating a cognitive deficit in amammal caused by brain damage, said method comprising administeringpluripotent cells into a damaged mammalian brain, wherein said damage isprimarily in one hemisphere and said administration is into the brainhemisphere contra-lateral to the brain hemisphere containing the primarysite of damage, wherein said pluripotent cells are hippocampal mousecells comprising a gene encoding the tsA58 mutant of the SV40 large Tantigen under the control of an H-2K^(b) promoter, wherein saidpluripotent cells are capable of differentiating into neural cells, andwherein said pluripotent cells are administered in an amount effectiveto improve cognitive function.
 2. The method, according to claim 1,wherein brain damage is associated with stroke.
 3. The method, accordingto claim 1, wherein said pluripotent cells are neuroepithelial stemcells.
 4. The method, according to claim 1, wherein said pluripotentcells are from the MHP36 clonal cell line.
 5. The method, according toclaim 1, wherein the brain is stimulated by exercise after administeringthe pluripotent cells.
 6. The method, according to claim 5, wherein saidexercise is augmented by bio-feedback.
 7. The method, according to claim1, wherein said pluripotent cells are administered only into the brainhemisphere contra-lateral to the brain hemisphere containing the primarysite of damage.
 8. The method, according to claim 1, wherein saidpluripotent cells are administered at multiple sites into the brainhemisphere contra-lateral to the brain hemisphere containing the primarysite of damage.
 9. The method, according to claim 1, wherein saidpluripotent cells are administered into an undamaged area of the brain.10. The method, according to claim 1, wherein said pluripotent cells areadministered intraparenchymally.
 11. The method, according to claim 1,wherein said pluripotent cells are administered intraparenchymally intoan undamaged area of the brain.
 12. The method, according to claim 7,wherein said pluripotent cells are administered into an undamaged areaof the brain.
 13. The method, according to claim 7, wherein saidpluripotent cells are administered intraparenchymally.
 14. The method,according to claim 7, wherein said pluripotent cells are administeredintraparenchymally into an undamaged area of the brain.
 15. The method,according to claim 1, wherein said pluripotent cells are derived from anH-2K^(b)-tsA58 transgenic mouse.